V. Molony

An α2 receptor mediates the selective inhibition by noradrenaline of nociceptive responses of identified dorsal horn neurones

Extracellular recordings were made of 59 neurones with long, ascending projections (spinocervical tract (SCT) and dorsal column postsynaptic (DCPS) neurones) in the lumbar dorsal horn of anaesthetized and paralyzed cats. All showed prominent excitatory responses to innocuous stimuli, applied to their cutaneous receptive fields on the ipsilateral hindlimb. The majority of the population investigated (83%) was multireceptive, being activated by noxious as well as innocuous cutaneous stimuli. Drug effects were examined on a regular cycle of responses to these cutaneous stimuli and also to DL-homocysteic acid (DLH). In 49 multireceptive SCT and DCPS neurones, ionophoretically-applied L-noradrenaline (NA) produced a potent selective inhibition of the nociceptive responses (to heat or pinch) in 40 out of 44 SCT and 3 out of 5 DCPS neurones, with no statistically significant change in the responses to innocuous brush or DLH, or in spontaneous activity. NA had no effect on the majority of cells (8 out of 11) that responded only to innocuous stimuli. In 19 SCT neurones that showed NA-selectivity, the alpha 2-selective agonists clonidine (in 12 out of 15) and metaraminol (in 2 out of 3) mimicked this selective effect, whereas, the alpha 1 agonist, phenylephrine and the beta agonist, isoprenaline did not. Furthermore, the alpha 2 antagonists, yohimbine and idazoxan (RX781094), either reversed or reduced the potency of the NA-elicited inhibition of nociceptive responses in all 7 SCT neurones tested. These results are discussed in relation to other evidence for spinal antinociceptive effects of noradrenergic systems acting at a spinal level and the possible involvement of an alpha 2 receptor in such effects.

The involvement of neurokinin receptor subtypes in somatosensory processing in the superficial dorsal horn of the cat

As well as substance P (SP), neurokinin A (NKA) and neurokinin B (NKB) have recently been found in the superficial dorsal horn of the spinal cord; NKA originating mainly in fine primary afferents. We have investigated the effects of these tachykinins and a range of analogues on somatosensory responses of single identified dorsal horn neurons, when applied ionophoretically to the region of the substantia gelatinosa. Behavioural reflex tests of thermal nociception were carried out in parallel. The role of NK-1, NK-2 and NK-3 receptors was addressed. NK-1-selective agonists attenuated the non-nociceptive responses of identified multireceptive spinocervical tract (SCT) neurons. Of the endogenous tachykinins, both SP and NKB (a weak NK-1 agonist) showed this effect. No role for NK-3 receptors was identified in our experiments. NK-2-selective agonists (including NKA) caused a unique and selective facilitation of thermal nociceptive responses. NKA also reduced reflex response latency in tail-flick and hot plate tests. NKA as a primary afferent transmitter may thus be involved in mediating or facilitating the expression of thermal nociceptive inputs in the substantia gelatinosa. NKA and SP could be considered as acting in concert in the superficial dorsal horn in an effectively pro-nociceptive modulatory role. Evidence from receptor-selective antagonists supports that obtained with agonists for the roles of particular NK receptors in somatosensory processing. NK-2, but not NK-1 or NK-3 antagonists attenuated endogenous thermal nociceptive responses, supporting the hypothesis that an NK-2 agonist (such as NKA) may normally participate in expression of thermal nociception in the superficial dorsal horn. Behavioural experiments showing increased response latencies with a putative NK-2 selective antagonist further supported the involvement of NK-2 receptors in thermal nociception.

Responses of spinocervical tract neurones to noxious stimulation of the skin.

1. Activity of single spinocervical tract neurones has been recorded in the lumbar spinal cord of chloralose anaesthetized or decerebrated cats. Reversible spinalization was produced by cold block at L3. Sensitivity of these neurones to noxious stimulation was studied by heating their cutaneous receptive fields above 40‐45 degrees C. 2. Most of the units were located in lamina IV of the dorsal horn and had their receptive fields in the ipsilateral foot. All but one of the studied neurones were excited by moving hairs or by gentle mechanical stimulation of the skin. 3. Eighty‐four % of the units were affected by noxious stimuli and three kinds of response were obtained: (i) 61% were excited (E‐cells) by noxious heat; (ii) 19% were inhibited (I‐cells); and (iii) 19% gave a mixed response reversing from excitatory to inhibitory (EI‐cells). 4. E‐cells had axons with a wider range of conduction velocities than the rest and also received the strongest descending inhibition from supraspinal structures. 5. The recording sites of EI‐cells were located in the medial third of the dorsal horn whereas E‐ and I‐cells were distributed over the full width of the dorsal horn. 6. The possible role of the spinocervical tract in nociception is discussed.

Comparison of methods for the reduction of acute painproduced by rubber ring castration or tail docking of week-old lambs

Behavioural and plasma cortisol changes were recorded for groups of eight Suffolk x Greyface lambs subjected to castration or tail docking using rubber rings with and without local anaesthetic treatment. Immediately after application of the rubber ring, local anaesthetic (2 x 0.2 ml 2% lignocaine) was administered either by needle and syringe or by high-pressure needleless injection into each side of the neck of the scrotum or tail at the site of the ring, or by high pressure needleless injection into the testes before ring application. In other groups, the innervation to the scrotum or tail was disabled by crushing with a powered bloodless castrator just proximal to the ring. Measurements were recorded in groups of control (handled) lambs, with and without local anaesthetic treatment. Application of local anaesthetic by high pressure needleless injection had little effect on either plasma cortisol values or behaviour of control lambs. For castration, application of the bloodless castrator and/or local anaesthetic at the ring site reduced the peak plasma cortisol concentration by 50% (P < or = 0.01), the incidence of active behavioural responses by 80 and 64% (P < or = 0.01) respectively, and the time spent in abnormal postures by 68 and 59% (P < or = 0.01) respectively. Both methods were effective in reducing pain for tail docking [active behavioural responses reduced by more than 80% (P < or = 0.01) and abnormal postures by 56% (P < or = 0.01)], although local anaesthetic was more effective, reducing the peak cortisol by 60% (P < or = 0.01) [crushing by 44% (P > 0.05)]. Injection of local anaesthetic into the testes was less effective than injection into the neck of the scrotum at the site of the ring [reduction in abnormal lying postures (P < or = 0.05), 45 vs 71%, respectively]. The rapid action, effectiveness, and ease of application of these experimental methods may provide the basis for commercially viable methods for reducing the acute pain produced by rubber ring castration and tail docking of lambs.

The influence of opioid receptor subtypes on the processing of nociceptive inputs in the spinal dorsal horn of the cat.

Extracellular recordings were made of the cutaneous sensory responses of spinocervical tract (SCT) neurones in the lumbar dorsal horn of anaesthetised and paralysed cats. All of the neurones studied were multireceptive, showing excitatory responses to both innocuous and noxious (thermal and when tested, mechanical) stimuli applied to their cutaneous receptive fields on the ipsilateral hindlimb. The effects of iontophoretically applied opioids were studied on a regular cycle of responses to these cutaneous stimuli and also to D.L-homocysteic acid (DLH). In the first series of experiments, drugs were applied in the vicinity of the SCT neurones. The kappa-receptor agonists dynorphin A(1-13) and U50488H, but not dynorphin A(2-13), the mu-agonist DAGO, or the delta-agonist DADL, caused a selective reduction of the nociceptive responses of the neurones. The corresponding responses to innocuous stimuli or to DLH, and spontaneous activity were unaffected. In the second series of experiments, drugs were applied from a second electrode placed in the region of the substantia gelatinosa directly dorsal to the tip of the recording electrode. Under these conditions, the mu-receptor agonist DAGO, but not the kappa-agonist dynorphin A(1-13) or the delta-agonists DADL, DSLET or DLPEN, showed a selective antinociceptive effect. In both series, the antinociceptive effects of the opioids were readily reversed by iontophoretically applied naloxone. The effect of dynorphin A(1-13) applied close to SCT neurones, but not that of DAGO applied in the region of the substantia gelatinosa, was reversed by the alpha 2-adrenoceptor antagonist, idazoxan. The results indicate that both mu- and kappa-opioid receptors (at anatomically distinct sites) can participate in the selective antinociceptive influence that opioids can exert over somatosensory information ascending to supraspinal levels. The antagonism of kappa-receptor-mediated antinociception by idazoxan is consistent with an interaction of opioid and noradrenaline influences at the level of the dorsal horn.

Ascending projections of nociceptor-driven Lamina I neurones in the cat

SummarySingle unit activity has been recorded from nociceptor-driven Lamina I neurones in the lumbar spinal cord of chloralose anaesthetized and gallamine paralysed cats. Ninety-four nociceptor-driven Lamina I neurones were identified by their superficial location in the dorsal horn and their ability to respond only to noxius stimulation of their cutaneous receptive fields. One-third of the Lamina I neurones responded only to noxious mechanical stimulation of the skin (Class 3a) und two-thirds responded to both mechanical and thermal noxious stimulation (Class 3b). Lissauers tract was stimulated electrically two and three segments rostral to the recording sites. Ninety percent of the neurones tested showed a post-synaptic excitation mediated by fibres conducting at a mean velocity of 5.2 m/s (range 0.9–13.3 m/s). It is concluded that Aδ and C afferent fibres running in Lissauers tract excite nociceptor-driven Lamina I neurones. Ninety-six percent of the neurones tested showed a long period of inhibition (100–200 ms) following stimulation of large afferent fibres in the dorsal column. This inhibition was increased when the intensity of stimulation recruited Lissauers tract fibres. Fifteen percent of the neurones tested were antidromically activated by Lissauers tract stimulation from up to 3 segments rostal to their origin. A further 18.5% were antidromically excited by stimulation of deeper tracts. The mean conduction velocity of the axons of these projecting neurones was 8.6 m/s (range 3.8–16.5 m/s) and thus are small myelinated axons. The Class 3b neurones exhibited a significantly lower conduction velocity (7.5±2.8 (S.D.) m/s) than the Class 3a neurones (10.7±3.7 (S.D.) m/s).It is concluded that at least two-thirds of the population of nociceptor-driven Lamina I neurones are segmental interneurones.

Nociceptive neurones in the superficial dorsal horn of cat lumbar spinal cord and their primary afferent inputs

SummaryThe morphology, background activity and responses to stimulation of primary afferent inputs of small neurones in the superficial dorsal horn which could only be excited from the skin by noxious stimulation were investigated by intracellular recording and ionophoresis of HRP. Neurones which gave similar responses to afferent stimulation were morphologically heterogeneous with respect to dendritic tree geometry and axonal projection, but were located around the lamina I/II border. Cutaneous excitatory receptive fields responding to noxious stimulation were generally small; most neurones had more extensive inhibitory fields responding to innocuous mechanical stimulation, in many cases overlapping the excitatory fields. Generally, stimulation of the excitatory field resulted in depolarization of the neurone and increased action potential firing, and stimulation of the inhibitory field resulted in hyperpolarization. Electrical stimulation of peripheral nerves revealed the existence of converging excitatory inputs carried by different fibre groups, and all neurones received an inhibitory input activated at low threshold. Excitatory responses were short-lived and occurred consistently in response to repeated stimulation. Central delay measurements gave evidence of a number of Aδ monosynaptic inputs but only one Aβ monosynaptic input; inhibitory inputs along Aβ fibres were polysynaptic. The constant latency and regularity of the C response suggested monosynaptic connections. Low intensity stimulation of inhibitory inputs elicited a short-lived i.p.s.p. which increased in amplitude with increasing stimulus strength until it disappeared into a more prolonged hyperpolarization. This was associated with inhibition of background action potentials, and increased in duration with increasing stimulus strength up to C levels, indicating an Aδ and C component. It is suggested that the level of excitability of these neurones depends on the relative amounts of concurrent noxious and innocuous stimulation, and that the resultant output, which is conveyed mainly to other neurones within the spinal cord, could modulate reflex action at the spinal level as well as affecting components of ascending sensory pathways.

Castration of calves: a study of methods used by farmers in the United Kingdom

A postal survey of farmers was conducted to determine the main methods used to castrate calves, and by whom and how they were applied. Among the 28 per cent of farmers who replied, those who did castrate calves used one or more of three methods: the Burdizzo was used by 43 per cent of farmers, surgery by 39 per cent, and rubber rings by 32 per cent, with 10 per cent using more than one method. Calves were castrated at all ages from less than one week to over six months, with one third of them being castrated at an age that legally requires the operation to be done under local anaesthesia by a veterinary surgeon . Rubber rings were never used by veterinary surgeons, but they carried out 43 per cent of surgical castrations, which was the method of choice in older calves. Local anaesthetic was used on 15 per cent of farms, mainly for surgical castrations. Sixty-seven per cent of farmers using the Burdizzo applied it twice, with the majority correctly applying the second crush below the first, and 90 per cent used precautions to control infection after surgical castration.

Supraspinal linkage gelatinosa neurones: effects of descending impulses

It is now well established that sensory transmission through the dorsal horn can be modified by descending influences from supraspinal structures (for review see ref. 15). Most dorsal horn neurones in laminae IV, V and VI of Rexed 23 are under tonic descending inhibition14,18,19, 24, which particularly affects their responses to noxious inputs is. The specific nociceptor-driven neurones in lamina I are also under tonic descending inhibition from supraspinal centres s. Similar inhibitory actions have been described on spinal cord neurones projecting through the spinocervical tract4,5, 9, the spinothalamic tractZ,el, 27, and the spinoreticular tract 16. Several descending pathways, including the pyramidal tract 6, have been proposed as mediators of this descending control. Anatomical and functional studies have shown that most of these descending tracts course in the dorsolateral funiculus (DLF) of the spinal cord1,5,17 and terminate in the centre and base of the dorsal horn, i.e. laminae IV, V and VI. Basbaum et al. 2 have recently described three bulbospinal pathways from the rostral medulla of the cat, one of which originates in the nucleus raphe magnus and projects bilaterally through the DLF to laminae V, VI and VII and in addition to the most superficial dorsal horn, lamina I and the substantia gelatinosa (SG). The functional organization of the small neurones of the SG is now under intensive studyl0-~2,~0, ~5. In this paper we report results from a study on lumbar SG neurones of influences descending from more rostral parts of the central nervous system. Results have been obtained from cats anaesthetized with chloralose (60 mg/kg) and paralyzed with gallamine triethiodide, using standard procedures to ensure full anaesthesia of the animals at all times. SG neurones were recorded in the L6 and L7 segments of the cord using high impedance glass microelectrodes 2e. Tonic descending influences from supraspinal centres were tested by placing a thermode on the L1 segment which permitted reversible spinalization of the animals by cold block s. Balltyped surface electrodes were placed on the dorsolateral funiculus at L2-L3 level and

Membrane properties of nociceptive neurones in lamina II of lumbar spinal cord in the cat.

1. Intracellular recordings were made from neurones in lamina II of the dorsal horn of the lumbar spinal cord of the cat, and the electrotonic responses to brief rectangular current pulses of up to 0.5 nA passed through the recording microelectrode measured. 2. The majority of penetrations were associated with input resistances lower than 70 M omega, low resting potentials (‐25 to ‐45 mV) and frequent firing of action potentials. Stable resting potentials of ‐50 to ‐75 mV were recorded in twenty neurones which exhibited continuous ongoing synaptic activity without action potentials. The threshold for action potential initiation was around ‐42 mV. The current‐voltage relationships were linear over most of the range of currents used; with depolarizing currents rectification became apparent close to the firing threshold. Input resistances ranged from 80 to 150 M omega. 3. The time course of the decay of the electrotonic response was exponential with a time constant of 0.8‐2.0 ms. The morphology of the cells‐‐small soma with a small number of fine processes‐‐and these short time constants suggest that axial current flow is limited and that the charge is dissipated locally within the soma through the membrane capacitance. 4. Effective membrane capacities were calculated from the estimated soma surface area of typical neurones in lamina II stained with HRP, and assuming a specific membrane conductance of 1.0 microF cm‐2 they ranged from 3.1 to 15.7 pF. Membrane capacities were calculated for the twenty neurones in this study from measurement of input resistance and time constants (6.4‐15.0 pF) and lay within this range. 5. Three neurones which had their electrical properties measured were also stained with horseradish peroxidase (HRP). Their specific membrane capacitances (1.1‐1.2 microF cm‐2) and specific resistances (0.9‐1.1 k omega cm2) were within the range of values measured for other neurones in the CNS. 6. The short time constants found for these neurones suggests that temporal summation of postsynaptic potentials evoked by short‐acting neurotransmitters will be limited. This may help to explain why action potentials arise singly from discrete, short‐lived EPSPs. There is anatomical evidence for multiple connections from terminal branches of A delta and C afferent fibres within the superficial dorsal horn; this suggests that spatial summation of EPSPs is a major factor in synaptic integration of some of the primary afferent inputs to these neurones.